Modulation of fluorouracil cytotoxicity by interferon-alpha and -gamma.
Because interferons (IFN)-alpha and -gamma individually have increased fluorouracil (FUra) cytotoxicity in several in vitro models, we studied the effects of FUra combined with IFN-alpha + gamma in HT29 colon cancer cells. A 96-hr exposure to IFN-alpha (500 units/ml) plus IFN-gamma (10 units/ml) and a 72-hr exposure to 0. 25-1 microM FUra (hr 24-96) inhibited cell growth and colony formation in an additive or more-than-additive fashion. When cells were exposed to IFN-alpha + gamma and FUra, free FdUMP levels became detectable, whereas [3H]FUra-RNA incorporation decreased. Exposure to IFN-alpha + gamma, FUra, or the combination decreased dTTP pools to 58%, 43%, and 17% of control, respectively. A marked increase in the dATP to dTTP ratio was seen with FUra with or without IFN-alpha + gamma. Thymidylate synthase catalytic activity was reduced to 28% and 24% of control with FUra with or without IFN-alpha + gamma, suggesting that the enhanced dTTP depletion must be due to another mechanism. FUra-mediated thymidylate synthase inhibition was accompanied by a 124-fold increase in total deoxyuridylate immunoreactivity and a 31-fold increase in dUTP pools, but the addition of IFN-alpha + gamma attenuated the accumulation. Treatment with IFN-alpha + gamma and FUra individually interfered with nascent DNA chain elongation, whereas the three-drug combination produced the most striking effects. IFN-alpha + gamma plus FUra produced the greatest amount of single-strand breaks in nascent DNA and dramatically decreased net DNA synthesis. IFN-alpha + gamma with or without FUra produced double-strand breaks in parental DNA. These results suggest that dTTP depletion, dATP/dTTP imbalance, pronounced inhibition of DNA synthesis, and damage to nascent and parental DNA contribute to the enhanced cytotoxicity with the triple combination.
Ismail
A
A
National Cancer Institute-Navy Medical Oncology Branch, National Naval Medical Center, Bethesda, Maryland 20889, USA.
Van Groeningen
C J
CJ
Hardcastle
A
A
Ren
Q
Q
Aherne
G W
GW
Geoffroy
F
F
Allegra
C J
CJ
Grem
J L
JL
D053938:DNA Fragmentation; D004260:DNA Repair / Q000187:drug effects; D004273:DNA, Neoplasm / Q000096:biosynthesis; D004334:Drug Administration Schedule; D004347:Drug Interactions; D005472:Fluorouracil / Q000008:administration & dosage* / Q000378:metabolism; D006801:Humans; D016898:Interferon-alpha / Q000008:administration & dosage*; D007371:Interferon-gamma / Q000008:administration & dosage*; D012334:RNA, Neoplasm / Q000096:biosynthesis; D013942:Thymine Nucleotides / Q000378:metabolism; D014407:Tumor Cells, Cultured / Q000187:drug effects
D004273:DNA, Neoplasm; D016898:Interferon-alpha; D012334:RNA, Neoplasm; D013942:Thymine Nucleotides; D007371:Interferon-gamma; C024157:thymidine 5'-triphosphate; D005472:Fluorouracil